1. Identify the sources of somatic cell lineages in the fetal gonads and investigate how they acquire their organ-specific identities This project provides the first genetic evidence for the origins of theca progenitor cells and their lineage commitment in the ovary. Theca progenitor cells from divergent sources (mesonephros and somatic precursors of the fetal ovary) acquire the expression of Gli1 during their commitment to the theca cell lineage. The expression of Gli1 in theca progenitor cells is indirectly induced by the oocyte-specific factor GDF9 through Hh ligands (DHH and IHH) in granulosa cells (see Model on the right). In the absence of Hedgehog signaling in the ovary, theca progenitor cells fail to develop, leading to defects in folliculogenesis and ovulation. The findings of this project raise some interesting questions and open new research directions. For instance, what is the source of signal(s) that trigger the migration of the Gli1-positive cells from the mesonephros? What is the functional significance of mesonephros-derived and ovary-derived theca cells? We have developed plans to answer these questions in the future (see next section). The topic of theca cell development is fundamentally important, given that disorders in theca cell differentiation are implicated in ovarian diseases such as polycystic ovary syndrome (PCOS), premature ovarian failure, and ovarian cancers. The discovery of the origins of theca cells and the mechanism underlying their development serve as novel entry points to understanding how theca cell-related pathology affects female reproductive health. 2. Define the cellular and molecular processes that lead to sexually dimorphic establishment of the reproductive tracts Before sexual differentiation occurs, embryos are anatomically bisexual as they possess both male and female reproductive tracts. These two tracts derive from two separate progenitor systems in the fetal mesonephros: Wolffian duct for the male tract and Mllerian duct for the female tract. In the male embryos, Wolffian ducts are maintained by testis-derived androgens while Mllerian duct undergo regression induced by anti-Mllerian hormone, also a product of testes. Female embryos, which do not produce androgens or anti-Mllerian hormone (AMH), experience the opposite where Mllerian ducts are maintained and Wolffian ducts undergo regression. In search of novel regulators in this process, we discovered the presence of orphan nuclear receptor COUPTFII in the mesenchyme of the mesonephros. Inactivation of COUPTFII specifically in the mesonephric mesenchyme leads to maintenance of both Wolffian and Mllerian ducts in the male and female mouse embryos, a typical case of pseudohermaphroditism. The affected embryos still have sex-specific production of hormones (androgen and AMH), indicating that hormonal abnormality is not responsible for the pseudohermaphroditic phenotypes of the reproductive tracts. Instead, COUPTFII in the mesenchyme of the mesonephros appears to be a molecular switch that controls the decision-making process of the identity of reproductive tract progenitors. This novel finding provides new mechanistic insights into the dimorphic establishment of reproductive tract. Ongoing experiments are to identify the upstream and downstream regulators of COUPTFII in this process. 3. Investigate the effects of in utero exposure to endocrine disruptors on the development of fetal reproductive organs and its lingering impacts on fertility in adulthood Formation of fetal reproductive organs relies on an intricate interaction between steroid hormones and signaling molecules, therefore making this process a prime target of endocrine disruptors. Chemicals or compounds that mimic or interfere with the action of steroid hormone and signaling molecules are known to have detrimental impacts on fetal reproductive organ formation and long-term impacts on fertility when the affected animals reach adulthood. Arsenic, a human carcinogen found in underground water and food products, has been shown to affect the Hedgehog pathway, one of the signaling pathways important for the formation of reproductive organs. Exposure of moues embryos to arsenic leads to cancer development in the ovary and reproductive tracts, organs where the Hedgehog pathway is functional. To investigate whether arsenic exposure targets the Hedgehog pathway in the fetal gonads and reproductive tract, we expose pregnant mice with various doses of inorganic arsenic in the drinking water. We found a dose-dependent and organ-specific effect of arsenic exposure on the Hh pathway: arsenic activates the Hedgehog pathway in the ovary whereas decrease its activity in the fetal testis. When some of the exposed female fetuses are allowed to develop to adulthood, they exhibit precocious onset of puberty (vaginal opening) compared to the control littermate. The Hedgehog pathway is known to control steroidogenesis in the ovary and testis. We therefore hypothesize that altered Hedgehog activity by in utero arsenic exposure could contribute to premature steroid (i.e. estrogen) production and consequent early onset of puberty in the affected female. Our findings are the first to demonstrate a potential impact of arsenic exposure on female reproductive systems. We are collaborating with Jean Harry and Mike Walkkes at NTP to study the neurological impacts of arsenic on these animals.